AZENTA NGS DATA REPORT

1. Project Summary

Project ID:30-775111958
Customer:SeanOConnell
BI:GENEWIZ-NGS-BI

2. Description of Workflow

2.1. Experiment Workflow

16S/18S/ITS rRNA is composed of conserved and hypervariable regions. Whereas conserved regions are not significantly different across various microbial strains, the sequences of hypervariable regions are genus or species-specific, and differ in accordance to phylogenetic difference. Therefore, 16S/18S/ITS rDNA serve as identifiers of biological species, and are important for microbial phylogeny and taxonomic identification. 16S/18S/ITS rDNA amplicon sequencing has become an important tool for the study of the composition of microbial communities in environment.

16S rDNA amplicon sequencing includes the library construction using specific primers to amplify the variable region of prokaryotic 16S rDNA and data analysis of the 16S rDNA variable region sequence to identify the composition and abundance of prokaryotic microorganisms in the environment. The proprietary workflow at Azenta effectively amplifies the three variable regions of 16S rDNA (V3, V4, V5) and accurately identifies various species including archaea. For samples with eukaryotic contamination, only V3, V4 region are amplified. 18S / ITS rDNA amplicon sequencing includes the library construction using specific primers to amplify the variable region of eukaryotic 18S / ITS rDNA and data analysis to identify the composition and abundance of eukaryotic microorganisms in the environment. Illumina MiSeq sequencing platform is widely used for 16S / 18S / ITS rDNA amplicon sequencing because of its deep sequencing depth, high throughput, short run-time and high sequencing accuracy as well as reasonable cost. In recent years, pair-end chemistry has enabled MiSeq sequencing platform to read as long as 600bp, which further increased the accuracy of the results.

16S / 18S / ITS rDNA amplicon sequencing procedure includes genomic DNA extraction, quality control, rDNA variable region amplification, library construction, high-throughput sequencing and data analysis. All the steps are important for data quality and quantity, which in turn affects the subsequent data analysis. In order to ensure data accuracy and reliability, every step has to pass strict quality control before pooling the library by adjusting the volume of each library according to the target data volume for Illumina MiSeq sequencing. The workflow is as below:

Figure 2.1 Microflora diversity experimental workflow

2.2. Bioinformatics Analysis Workflow

Figure 2.2 Microflora diversity data analysis workflow

Note:

(1) If the number of samples is less than 3, comparative analysis of diversity cannot be performed.

(2) The differential statistical analysis is meaningful only if there are at least three replicates in the biology group.

(3) Environmental factors are required for correlation analysis of community composition and environmental factors as well as for CCA / RDA analysis.

(4) Intestinal-type analysis only applies to animal or human intestinal or stool samples.

(5) Analysis with * are not included in the standard analysis and can be selected and analyzed according to individual sample and project.

3. Analysis

3.1. Sample Summary Statistics

SampleLength(bp)#ReadsBases(bp)Q20(%)Q30(%)GC(%)N(ppm)
C1241.0370,29016,942,09196.5891.1046.15141.19
C2-2246.75239,09258,995,50496.8891.4642.8532.90
C3248.29241,76260,028,27596.9291.5549.6243.16
P1246.83241,78059,679,34397.1091.8551.7231.30
P2244.71240,71058,903,76096.8491.5552.4428.44
P3247.37240,58459,512,64497.0991.8350.2732.87

3.2. Sequencing Data Quality Optimization

Sequencing errors might occur in high-throughput sequencing, and it is common that bases toward the end of the sequence reads have lower than average quality. In order to obtain higher quality and more accurate bioinformatic analysis results, it is necessary to optimize the raw data of the sequencing to obtain higher quality and more accurate bioinformatics analysis results.

Analysis software: Cutadapt(v1.9.1), Vsearch(1.9.6), Qiime(1.9.1)

Steps and parameters for optimization:

(1) The two sequences of each read pair were merged according to overlapping sequences. The read merge is deemed to be successful only if the overlapping sequence is least 20bp long. After merging, undetermined bases (N) were removed from the resulting sequence.

(2) Primer and adapter sequences were removed. Then the 5' and 3' bases with Q score lower than 20 were also removed. The resulting sequences with length > 200bp would pass this step of processing.

(3) The sequences obtained were then aligned to UCHIME 'Gold' database to identify and remove chimera sequence. Sequences passed this filtering step are deemed as clean data ready for analysis.

Sample#PE_reads#NochimeraAvgLen(bp)GC(%)Effective(%)
C135,14530,086326.3245.1885.61
C2_2119,546113,317332.7542.8194.79
C3120,881115,192339.2949.1295.29
P1120,890113,295332.6450.9393.72
P2120,355105,574323.8251.3687.72
P3120,292113,943339.9449.8694.72

Figure 3.2 Effective sequences length distribution

3.3. OTU Analysis and Species Annotation

3.3.1. OTU clustering

OTU is an operational definition of a classification unit (genus, species, grouping, etc.) commonly used in population genetics to facilitate data analysis. In bioinformatics each sequence obtained from sequencing is assumed to be derived from a single species. All the sequences in a sample are classified to obtain information on species and genus. By classification, the sequences are grouped according to their similarity, and one group is an OTU. Typically, OTU clusters are defined by a 97% identity threshold for data statistics and analysis.

Analysis software:Qiime(1.9.1),Vsearch(1.9.6)

Analysis methods and steps:

(1) Unique sequences were extracted from the optimized sequences with the read count information.

(2) The unique sequences with 1 read count were removed.

(3) OTU clustering of unique sequences (read count > 1) was performed with similarity of 97%, and chimeric sequences were further removed to obtain the representative OTU sequences.

(4) All optimized sequences were compared with OTU representative sequences, and sequences of >97% similarity to a specific OTU representative sequence are considered to be of the same OTU. Finally, the OTU abundances were also summarized in the results table.

The following table shows the statistics of the sequence number in each sample's OTU:

OTU_IDC1C2-2C3P1P2P3taxonomy
OTU417,96610,0061753292311k__Fungi;p__Mucoromycota;c__Mucoromycetes;o__Mucorales;f__Mucoraceae;g__Mucor;s__Mucor_circinelloides
OTU5459,258161120k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Tricholomataceae;g__Pseudotricholoma;s__Pseudotricholoma_umbrosum
OTU7001,18721328,029k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Russula
OTU8214825323,8032,9351,550k__Fungi;p__Basidiomycota;c__Tremellomycetes;o__Tremellales;f__Trimorphomycetaceae;g__Saitozyma;s__Saitozyma_podzolica
OTU60053,1052,3051,745k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Sebacinales;f__Sebacinaceae;g__Sebacina;s__unidentified
OTU963871,5363,08546725k__Fungi;p__Basidiomycota;c__Geminibasidiomycetes;o__Geminibasidiales;f__Geminibasidiaceae;g__Geminibasidium;s__unidentified
OTU140011,1272,4256k__Fungi;p__Ascomycota;c__Pezizomycetes;o__Pezizales;f__Pezizaceae
OTU111,0583111,682254335360k__Fungi;p__Ascomycota
OTU1227241952051,5681,310k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Leotiomycetes_ord_Incertae_sedis;f__Leotiomycetes_fam_Incertae_sedis;g__Leohumicola;s__Leohumicola_verrucosa
OTU100032,70997296k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Lactarius
OTU150012,1488187k__Fungi;p__Ascomycota
OTU16167671,681366401k__Fungi;p__Ascomycota
OTU1319321114771,591754k__Fungi;p__Ascomycota;c__Leotiomycetes
OTU28002,6427126k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Boletales;f__Boletaceae;g__unidentified;s__unidentified
OTU190021262,364k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Russula;s__Russula_crustosa
OTU31544891,673100242k__Plantae;p__unidentified;c__unidentified;o__unidentified;f__unidentified;g__unidentified;s__unidentified
OTU22001772,4523k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Boletales;f__Sclerodermataceae;g__Scleroderma;s__Scleroderma_citrinum
OTU17190932,3176k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Trichocomaceae;g__Sagenomella;s__Sagenomella_verticillata
OTU180032,05122k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Elaphomycetaceae;g__Elaphomyces;s__Elaphomyces_maculatus
OTU2370911701,192668k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Trichocomaceae;g__Sagenomella;s__Sagenomella_striatispora
OTU440047414505948k__Fungi;p__Ascomycota
OTU26101,499372230k__Fungi;p__Basidiomycota;c__Agaricomycetes
OTU2400011,9461k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Russula;s__Russula_cyanoxantha
OTU2500232541,512k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Sebacinales;f__Sebacinaceae;g__Sebacina;s__unidentified
OTU300008868641k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Russula;s__Russula_melzeri
OTU27121,58029000k__Fungi;p__Basidiomycota;c__Agaricomycetes
OTU200041,097346130k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Boletales;f__Astraeaceae;g__Astraeus;s__Astraeus_hygrometricus
OTU390028203410k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Sebacinales;f__Sebacinaceae;g__Sebacina;s__unidentified
OTU42749108130195273365k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Leotiomycetes_ord_Incertae_sedis;f__Leotiomycetes_fam_Incertae_sedis;g__Leohumicola;s__unidentified
OTU33001,062250k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Hygrophoraceae
OTU3231212114779735k__Fungi;p__Ascomycota
OTU293,60021010k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Elaphomycetaceae;g__Elaphomyces;s__unidentified
OTU3549400004k__Fungi;p__Ascomycota;c__Geoglossomycetes;o__Geoglossales;f__Geoglossaceae;g__Trichoglossum;s__Trichoglossum_walteri
OTU3630808411k__Fungi;p__Basidiomycota
OTU412027010843535k__Fungi;p__Mucoromycota;c__Endogonomycetes;o__GS20;f__unidentified;g__unidentified;s__unidentified
OTU4000176654112k__Fungi;p__Ascomycota
OTU21,3344640432k__Fungi
OTU521176470022k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Russula
OTU64003213102232k__Fungi;p__Ascomycota
OTU4830572251k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Clavariaceae;g__Clavulinopsis;s__Clavulinopsis_laeticolor
OTU4711363378797k__Fungi;p__Ascomycota;c__Saccharomycetes;o__Saccharomycetales;f__unidentified;g__unidentified;s__unidentified
OTU109203005763121k__Fungi;p__Ascomycota
OTU450012293236k__Fungi;p__Ascomycota;c__Pezizomycetes;o__Pezizales;f__Pezizaceae;g__Pachyphloeus;s__unidentified
OTU4900021457k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Thelephorales;f__Thelephoraceae
OTU5800446123k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Russula;s__Russula_rubellipes
OTU772869906415281k__Fungi;p__Ascomycota;c__Leotiomycetes
OTU5340463220k__Fungi;p__Ascomycota
OTU540002462431k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Inocybaceae;g__Inocybe;s__Inocybe_calamistrata
OTU7300405020k__Fungi;p__Ascomycota;c__Pezizomycetes;o__Pezizales;f__Pezizaceae;g__Hydnobolites
OTU48742252030166k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Leotiomycetes_ord_Incertae_sedis;f__Leotiomycetes_fam_Incertae_sedis;g__Leohumicola;s__Leohumicola_verrucosa
OTU600012410644154k__Fungi;p__Basidiomycota;c__Pucciniomycotina_cls_Incertae_sedis;o__GS25;f__unidentified;g__unidentified;s__unidentified
OTU430002617273k__Fungi;p__Ascomycota
OTU7400022387k__Fungi;p__Ascomycota
OTU7000119424651k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__unidentified;f__unidentified;g__unidentified;s__unidentified
OTU82113345457207k__Fungi;p__Ascomycota
OTU51013188995539k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella
OTU7223730000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Russula
OTU6200341120k__Fungi;p__Ascomycota
OTU5707344110k__Fungi;p__Ascomycota;c__Pezizomycetes;o__Pezizales;f__Tuberaceae;g__Tuber
OTU5612239591316k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Hypocreaceae;g__Trichoderma
OTU6100033043k__Plantae
OTU16241730000k__Fungi
OTU7176123518181k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Aspergillaceae;g__Penicillium
OTU6613080000k__Fungi;p__Ascomycota
OTU63617179185039k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales
OTU1490001720171k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Boletales;f__Boletaceae;g__Tylopilus;s__Tylopilus_atronicotianus
OTU7833112115888k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Nectriaceae;g__Ilyonectria;s__Ilyonectria_radicicola
OTU4693131551355k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Bondarzewiaceae;g__Bondarzewia;s__Bondarzewia_mesenterica
OTU84301321411140k__Fungi;p__Ascomycota
OTU12912544220711k__Fungi;p__Ascomycota;c__Dothideomycetes;o__Dothideomycetes_ord_Incertae_sedis;f__Gloniaceae;g__Cenococcum
OTU760022628287k__Fungi;p__Ascomycota
OTU6800246101k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Lactarius;s__Lactarius_chrysorrheus
OTU5952411743120k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella;s__Mortierella_pseudozygospora
OTU94000563211k__Fungi;p__Ascomycota
OTU670120125010k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Hygrophoraceae;g__Cuphophyllus;s__Cuphophyllus_lacmus
OTU10600342117115k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Chaetosphaeriales;f__Chaetosphaeriaceae;g__Chaetosphaeria;s__unidentified
OTU9000239100k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Russula;s__Russula_crustosa
OTU6961756144796k__Fungi;p__Mucoromycota;c__Umbelopsidomycetes;o__Umbelopsidales;f__Umbelopsidaceae;g__Umbelopsis;s__Umbelopsis_dimorpha
OTU790015731735k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella
OTU83250325630105k__Fungi;p__Ascomycota;c__Ascomycota_cls_Incertae_sedis;o__Ascomycota_ord_Incertae_sedis;f__Ascomycota_fam_Incertae_sedis;g__Trichocladium;s__Trichocladium_opacum
OTU1500347286142k__Fungi;p__Ascomycota
OTU10002535157085k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Clavicipitaceae;g__Metarhizium;s__Metarhizium_carneum
OTU115190201220k__Fungi;p__Ascomycota
OTU9627213638104k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Leotiomycetes_ord_Incertae_sedis;f__Pseudeurotiaceae;g__Pseudogymnoascus;s__Pseudogymnoascus_roseus
OTU12200183020k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Lactarius;s__Lactarius_quietus
OTU113570801906k__Fungi;p__Ascomycota;c__Dothideomycetes;o__Dothideomycetes_ord_Incertae_sedis;f__Gloniaceae;g__Cenococcum
OTU159188171371k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella;s__Mortierella_humilis
OTU98116729000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Clavariaceae;g__Clavaria
OTU12500190030k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Atheliales;f__Atheliaceae;g__Byssocorticium;s__Byssocorticium_pulchrum
OTU81150317158k__Fungi;p__Ascomycota;c__Pezizomycotina_cls_Incertae_sedis;o__GS35;f__unidentified;g__unidentified;s__unidentified
OTU9202410527233k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Aspergillaceae;g__Penicillium;s__Penicillium_verhagenii
OTU15445132715012k__Plantae;p__Anthophyta;c__Monocotyledonae;o__Poales;f__Poaceae;g__unidentified;s__unidentified
OTU107209921161k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Onygenales;f__unidentified;g__unidentified;s__unidentified
OTU99048997612k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella;s__Mortierella_beljakovae
OTU104007645727k__Fungi;p__Ascomycota;c__Archaeorhizomycetes;o__unidentified;f__unidentified;g__unidentified;s__unidentified
OTU8000169020k__Fungi;p__Ascomycota
OTU12800010169k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Amanitaceae;g__Amanita;s__Amanita_alboflavescens
OTU126183157010k__Fungi;p__Ascomycota
OTU8821750001k__Fungi;p__Ascomycota
OTU897120423379k__Fungi;p__Ascomycota
OTU9700010148k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Nectriaceae;g__Gliocladiopsis;s__Gliocladiopsis_curvata
OTU2400013510100k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella
OTU9100154120k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Clavariaceae;g__Clavaria;s__Clavaria_falcata
OTU102230143120k__Fungi;p__Ascomycota
OTU10300137010k__Fungi;p__Ascomycota;c__Xylonomycetes;o__GS34;f__unidentified;g__unidentified;s__unidentified
OTU13403299903k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Thelephorales;f__Thelephoraceae;g__Tomentella
OTU9300145000k__Fungi;p__Ascomycota
OTU157617812247k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella;s__Mortierella_humilis
OTU11200025193k__Fungi
OTU8500171560k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Hypocreaceae;g__Trichoderma
OTU19100039068k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Russula;s__unidentified
OTU13800112001k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Clavariaceae;g__Clavulinopsis
OTU120115227829k__Fungi;p__Basidiomycota;c__Tremellomycetes;o__Filobasidiales;f__Piskurozymaceae;g__Solicoccozyma;s__Solicoccozyma_terricola
OTU145055992920k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella;s__Mortierella_horticola
OTU10501123000k__Fungi;p__Ascomycota
OTU121001137446k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Aspergillaceae;g__Penicillium;s__unidentified
OTU35011326218k__Fungi
OTU13321130000k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales;f__Dermateaceae
OTU14200112320k__Fungi;p__Ascomycota
OTU11000013800k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales;f__Helotiaceae;g__Cudoniella;s__Cudoniella_acicularis
OTU16500101010k__Fungi
OTU18400108535k__Fungi
OTU12300115100k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Boletales;f__Calostomataceae;g__Calostoma;s__Calostoma_cinnabarinum
OTU22890105110k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Sebacinales;f__Sebacinaceae;g__Sebacina;s__unidentified
OTU17700826075k__Fungi;p__Ascomycota
OTU186009384025k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Aspergillaceae;g__Penicillium
OTU9500011300k__Fungi;p__Ascomycota;c__Pezizomycetes;o__Pezizales;f__Pezizaceae
OTU173991938339k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Sordariales;f__Sordariales_fam_Incertae_sedis;g__Dendrosporium;s__unidentified
OTU2690010297k__Fungi
OTU2040000096k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Thelephorales;f__Thelephoraceae;g__Tomentella
OTU12470100025k__Fungi;p__Ascomycota
OTU114365110000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Russula;s__Russula_chloroides
OTU15200011101k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Lactifluus
OTU1640068677k__Fungi
OTU1010099010k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Amanitaceae;g__Amanita
OTU20600000105k__Fungi;p__Glomeromycota;c__Glomeromycetes;o__Glomerales;f__Glomeraceae;g__Rhizophagus;s__Rhizophagus_irregularis
OTU18111086010k__Fungi;p__Ascomycota
OTU250253212423k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales;f__Helotiales_fam_Incertae_sedis;g__Leptodontidium;s__Leptodontidium_trabinellum
OTU135015323414k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Leotiomycetes_ord_Incertae_sedis;f__Pseudeurotiaceae
OTU1321940000k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Leotiomycetes_ord_Incertae_sedis;f__Leotiomycetes_fam_Incertae_sedis;g__Leohumicola;s__unidentified
OTU1670068000k__Fungi
OTU1930009301k__Fungi;p__Basidiomycota
OTU19700014154k__Fungi;p__Ascomycota
OTU1110008440k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Russula;s__Russula_subvinosa
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OTU23312368020k__Fungi;p__Ascomycota
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OTU224003501717k__Fungi
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OTU442160401178k__Fungi
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OTU2300069100k__Fungi
OTU2220054021k__Fungi
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OTU1440059010k__Fungi;p__Ascomycota
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OTU1750051020k__Fungi
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OTU1740056112k__Fungi
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OTU1850148710k__Fungi;p__Ascomycota
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OTU2250048000k__Fungi;p__Ascomycota
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OTU1920002929k__Fungi;p__Ascomycota
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OTU38000513128k__Fungi;p__Ascomycota
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OTU1700038110k__Fungi;p__Ascomycota
OTU2510004200k__Fungi;p__Ascomycota
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OTU2620000143k__Fungi;p__Ascomycota
OTU3154039000k__Fungi;p__Ascomycota
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OTU12700018121k__Fungi
OTU2380040030k__Fungi;p__Ascomycota;c__Dothideomycetes;o__Dothideomycetes_ord_Incertae_sedis;f__Gloniaceae;g__Cenococcum
OTU1820039010k__Fungi;p__Ascomycota
OTU1680039000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Russula
OTU4645081823k__Fungi
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OTU3930000480k__Fungi
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OTU3720002872k__Fungi;p__Ascomycota
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OTU41300011123k__Fungi;p__Ascomycota
OTU3019721010k__Fungi;p__Ascomycota
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OTU2900916100k__Fungi;p__Glomeromycota
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OTU27571150015k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales
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OTU2310133000k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella;s__Mortierella_parvispora
OTU2420036000k__Fungi;p__Ascomycota
OTU29501814030k__Fungi;p__Ascomycota
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OTU3390042100k__Fungi;p__Ascomycota
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OTU3600124830k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Sordariales;f__Chaetomiaceae
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OTU33300210010k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Clavicipitaceae
OTU2050001025k__Fungi;p__Ascomycota
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OTU3360026005k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Lycoperdaceae;g__Lycoperdon
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OTU3220022000k__Fungi;p__Ascomycota
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OTU18700180160k__Fungi;p__Ascomycota;c__Pezizomycetes;o__Pezizales;f__Pezizaceae;g__unidentified;s__unidentified
OTU4172000311k__Fungi;p__Ascomycota
OTU3210020010k__Fungi
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OTU3374036000k__Fungi
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OTU2390022000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Hygrophoraceae
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OTU1721120000k__Fungi;p__Ascomycota
OTU3070027000k__Fungi
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OTU4454230000k__Fungi
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OTU3770002024k__Fungi
OTU2941273000k__Fungi;p__Ascomycota
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OTU2560021600k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Nectriaceae;g__Volutella;s__unidentified
OTU1790017000k__Fungi
OTU21502111000k__Fungi;p__Basidiomycota;c__Geminibasidiomycetes;o__Geminibasidiales;f__Geminibasidiaceae;g__Geminibasidium;s__unidentified
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OTU3300019000k__Fungi
OTU3870313105k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Hypocreaceae;g__Trichoderma
OTU5760024000k__Fungi
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OTU4030000230k__Fungi;p__Ascomycota
OTU3570020010k__Fungi
OTU5580022110k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella
OTU4950301323k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Clavicipitaceae
OTU5141380133k__Fungi;p__Ascomycota;c__Sordariomycetes
OTU4462564001k__Fungi
OTU2643030230k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Tricholomataceae;g__Delicatula;s__Delicatula_integrella
OTU3700002300k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Nectriaceae;g__Nectria;s__Nectria_ramulariae
OTU4414620312k__Fungi;p__Ascomycota;c__Dothideomycetes;o__Pleosporales;f__Didymellaceae;g__Neoascochyta;s__Neoascochyta_exitialis
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OTU5300019000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Clavariaceae;g__Ramariopsis;s__unidentified
OTU3190017010k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Cortinariaceae;g__Cortinarius
OTU3250017000k__Fungi;p__Ascomycota
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OTU6970000210k__Fungi;p__Ascomycota;c__Eurotiomycetes
OTU3350015000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Hygrophoraceae;g__Gliophorus;s__unidentified
OTU2432021000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Amanitaceae;g__Amanita;s__Amanita_constricta
OTU4050000115k__Fungi;p__Ascomycota;c__Dothideomycetes;o__Pleosporales
OTU2203414000k__Fungi;p__Ascomycota
OTU28221101234k__Fungi
OTU2895310140k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Aspergillaceae;g__Penicillium;s__unidentified
OTU4040000320k__Fungi;p__Ascomycota
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OTU3680001146k__Fungi;p__Ascomycota
OTU4140000210k__Fungi;p__Ascomycota
OTU2350017000k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Sordariales
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OTU2490015040k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales
OTU3790001710k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales
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OTU6500001412k__Fungi;p__Ascomycota;c__Dothideomycetes;o__Pleosporales
OTU2550001401k__Fungi;p__Ascomycota
OTU4310000014k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Cantharellales;f__Ceratobasidiaceae
OTU2170150000k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales
OTU2650000200k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Onygenales;f__Onygenaceae;g__unidentified;s__unidentified
OTU2450018000k__Fungi;p__Ascomycota;c__Geoglossomycetes;o__Geoglossomycetes_ord_Incertae_sedis;f__Geoglossomycetes_fam_Incertae_sedis;g__Nothomitra;s__Nothomitra_cinnamomea
OTU3480011000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Tremellodendropsidales;f__unidentified;g__unidentified;s__unidentified
OTU3090016000k__Fungi
OTU6850000119k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales;f__Helotiales_fam_Incertae_sedis;g__Leptodontidium;s__Leptodontidium_trabinellum
OTU4503820102k__Fungi
OTU6280001400k__Fungi;p__Ascomycota;c__Dothideomycetes;o__Pleosporales;f__Fenestellaceae;g__Fenestella;s__Fenestella_mackenziei
OTU4150061120k__Fungi;p__Ascomycota
OTU5490013000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Boletales;f__Boletaceae;g__Strobilomyces;s__Strobilomyces_strobilaceus
OTU3440018000k__Fungi;p__Basidiomycota
OTU3200017000k__Fungi;p__Mucoromycota;c__Mucoromycetes;o__Mucorales;f__Mucoraceae;g__Mucor;s__Mucor_silvaticus
OTU2260024000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Thelephorales;f__Thelephoraceae
OTU5200511003k__Fungi;p__Ascomycota
OTU1900016000k__Fungi
OTU3040012000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Atheliales;f__Atheliaceae;g__Byssocorticium;s__unidentified
OTU5420015000k__Fungi;p__Ascomycota
OTU3240014000k__Fungi;p__Glomeromycota
OTU2982471401k__Fungi;p__Basidiomycota;c__Microbotryomycetes;o__Sporidiobolales;f__Sporidiobolaceae
OTU3980000110k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Boletales;f__Melanogastraceae
OTU212260020k__Fungi
OTU4300000017k__Fungi;p__Ascomycota;c__Sordariomycetes
OTU4370000018k__Fungi;p__Mucoromycota;c__Umbelopsidomycetes;o__Umbelopsidales;f__Umbelopsidaceae;g__Umbelopsis
OTU5400021010k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Aspergillaceae;g__Penicillium;s__unidentified
OTU261111077k__Fungi;p__Mucoromycota;c__Umbelopsidomycetes;o__Umbelopsidales;f__Umbelopsidaceae;g__Umbelopsis;s__Umbelopsis_ovata
OTU602009000k__Fungi;p__Ascomycota
OTU5050170000k__Fungi;p__Ascomycota
OTU5520011101k__Fungi;p__Ascomycota;c__Orbiliomycetes;o__GS33;f__unidentified;g__unidentified;s__unidentified
OTU6694400010k__Fungi
OTU3230015001k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Inocybaceae;g__Inocybe
OTU517210254k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Sordariales;f__Chaetomiaceae;g__Chaetomium
OTU6840000120k__Fungi;p__Ascomycota;c__Pezizomycetes;o__Pezizales;f__Pezizaceae;g__Hydnobolites;s__unidentified
OTU5470017000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales
OTU3590016000k__Fungi
OTU570003052k__Fungi;p__Basidiomycota;c__Microbotryomycetes;o__unidentified;f__unidentified;g__unidentified;s__unidentified
OTU6130018000k__Fungi
OTU268000117k__Fungi;p__Ascomycota;c__Geoglossomycetes;o__Geoglossales;f__Geoglossaceae;g__Trichoglossum
OTU4290000313k__Fungi;p__Glomeromycota;c__Archaeosporomycetes;o__Archaeosporales;f__Archaeosporaceae;g__Archaeospora;s__unidentified
OTU3660001600k__Fungi
OTU2808014000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Cyphellaceae;g__Rectipilus;s__Rectipilus_idahoensis
OTU621000850k__Fungi
OTU353002181k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Russula;s__Russula_changbaiensis
OTU4240000312k__Fungi;p__Glomeromycota;c__Archaeosporomycetes;o__Archaeosporales;f__unidentified;g__unidentified;s__unidentified
OTU2190100000k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales
OTU572001405k__Fungi
OTU3280014000k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Xylariales;f__Sporocadaceae;g__Pestalotiopsis;s__unidentified
OTU5750014001k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Trechisporales;f__Hydnodontaceae;g__Trechispora
OTU2700000018k__Fungi;p__Basidiomycota;c__Tremellomycetes;o__Tremellales;f__Tremellaceae;g__Tremella;s__unidentified
OTU23615011010k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales;f__Dermateaceae;g__Rhizodermea;s__Rhizodermea_veluwensis
OTU456930231k__Fungi;p__Ascomycota;c__Dothideomycetes;o__Capnodiales
OTU3380012000k__Fungi
OTU4350000015k__Fungi;p__Glomeromycota;c__Glomeromycetes;o__Glomerales
OTU5590013010k__Fungi;p__Glomeromycota;c__Glomeromycetes;o__Glomerales;f__Glomeraceae
OTU573008010k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Amanitaceae;g__Amanita;s__Amanita_nivalis
OTU4080100130k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Polyporales;f__Ganodermataceae;g__Ganoderma;s__Ganoderma_lucidum
OTU3420014000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Clavariaceae;g__Camarophyllopsis;s__Camarophyllopsis_atrovelutina
OTU7310000015k__Fungi;p__Ascomycota;c__Saccharomycetes;o__Saccharomycetales;f__Phaffomycetaceae;g__Cyberlindnera;s__Cyberlindnera_maclurae
OTU3830001000k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Coniochaetales;f__Coniochaetaceae;g__Coniochaeta
OTU5610012000k__Fungi;p__Ascomycota
OTU6780001150k__Fungi;p__Ascomycota
OTU422070070k__Fungi;p__Basidiomycota;c__Malasseziomycetes;o__Malasseziales;f__Malasseziaceae;g__Malassezia;s__Malassezia_globosa
OTU3580017000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Sebacinales;f__Sebacinaceae;g__Sebacina;s__unidentified
OTU2843530000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Tricholomataceae;g__Mycena;s__Mycena_abramsii
OTU699012210k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Trichocomaceae;g__Talaromyces
OTU4070000130k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Boletales;f__Boletaceae
OTU5740010001k__Fungi;p__Ascomycota
OTU310007000k__Fungi
OTU4260000210k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales
OTU6720000150k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Bionectriaceae;g__Clonostachys
OTU732001007k__Fungi;p__Ascomycota
OTU5310015000k__Fungi;p__Ascomycota;c__Pezizomycetes;o__Pezizales;f__Pezizaceae
OTU5330013010k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Sebacinales;f__Sebacinaceae;g__Sebacina;s__unidentified
OTU3910001150k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales
OTU494090000k__Fungi;p__Ascomycota
OTU466409011k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Aspergillaceae;g__Penicillium
OTU4180000140k__Fungi;p__Ascomycota
OTU710000550k__Fungi
OTU433000005k__Fungi;p__Glomeromycota;c__Glomeromycetes
OTU5240011000k__Fungi;p__Ascomycota;c__Pezizomycetes;o__Pezizales;f__Discinaceae;g__Hydnotrya;s__unidentified
OTU3780001100k__Fungi;p__Glomeromycota;c__Glomeromycetes;o__Glomerales;f__Glomeraceae;g__Rhizophagus
OTU671000180k__Fungi;p__Basidiomycota;c__Geminibasidiomycetes;o__Geminibasidiales;f__Geminibasidiaceae;g__Geminibasidium;s__unidentified
OTU4340000011k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Hypocreales_fam_Incertae_sedis;g__Acremonium
OTU3310012000k__Fungi
OTU6522300300k__Fungi;p__Basidiomycota;c__Tremellomycetes;o__Tremellales;f__Bulleribasidiaceae;g__Dioszegia;s__Dioszegia_hungarica
OTU639000500k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella;s__Mortierella_turficola
OTU536007000k__Fungi;p__Ascomycota
OTU3340012000k__Fungi
OTU3460010000k__Fungi;p__Ascomycota;c__Eurotiomycetes
OTU5480010000k__Fungi
OTU521021272k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Onygenales;f__Gymnoascaceae;g__Leucothecium;s__Leucothecium_emdenii
OTU4402421000k__Fungi;p__Basidiomycota;c__Tremellomycetes;o__Tremellales;f__Bulleribasidiaceae;g__Dioszegia;s__Dioszegia_fristingensis
OTU580013021k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Aspergillaceae;g__Penicillium
OTU3160013000k__Fungi;p__Mucoromycota;c__Endogonomycetes;o__GS20;f__unidentified;g__unidentified;s__unidentified
OTU254000800k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella;s__unidentified
OTU546003003k__Fungi
OTU593008000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Amanitaceae;g__Amanita
OTU605012070k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Aspergillaceae;g__Penicillium
OTU635400704k__Fungi;p__Ascomycota
OTU5570013000k__Fungi;p__Basidiomycota;c__Agaricomycetes
OTU345317000k__Fungi;p__Ascomycota
OTU665000071k__Fungi;p__Glomeromycota;c__Archaeosporomycetes;o__Archaeosporales;f__unidentified;g__unidentified;s__unidentified
OTU4110000130k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Boletales;f__Sclerodermataceae;g__Scleroderma;s__Scleroderma_areolatum
OTU682000035k__Fungi;p__Glomeromycota;c__Glomeromycetes;o__Glomerales
OTU6320001111k__Fungi;p__Ascomycota
OTU641000500k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Cantharellales;f__Cantharellales_fam_Incertae_sedis;g__Sistotrema;s__Sistotrema_sernanderi
OTU395000080k__Fungi
OTU3500012000k__Fungi;p__Glomeromycota;c__Archaeosporomycetes;o__Archaeosporales;f__Ambisporaceae;g__Ambispora;s__unidentified
OTU349007000k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales
OTU402000060k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Albatrellaceae;g__Leucogaster
OTU341450000k__Fungi
OTU327009000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Thelephorales;f__Thelephoraceae
OTU375000090k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Russula;s__unidentified
OTU692000033k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales
OTU690000090k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales
OTU341006013k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella;s__Mortierella_parvispora
OTU376000800k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Clavariaceae;g__Clavaria
OTU743000007k__Fungi
OTU195000500k__Fungi;p__Ascomycota
OTU4100000150k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Hydnangiaceae;g__Laccaria
OTU259000080k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Sebacinales;f__Sebacinaceae;g__Sebacina;s__unidentified
OTU733000008k__Fungi;p__Glomeromycota;c__Archaeosporomycetes;o__Archaeosporales;f__Archaeosporaceae
OTU677000090k__Fungi
OTU312006000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Russula;s__Russula_flavida
OTU485491000k__Fungi
OTU539006010k__Fungi;p__Glomeromycota;c__Archaeosporomycetes;o__Archaeosporales;f__Archaeosporaceae
OTU438500004k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Sordariales;f__unidentified;g__unidentified;s__unidentified
OTU578108000k__Fungi;p__Ascomycota
OTU739000007k__Fungi;p__Ascomycota
OTU329008000k__Fungi;p__Ascomycota;c__Lecanoromycetes
OTU5660011000k__Fungi;p__Ascomycota
OTU726000008k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella;s__Mortierella_hyalina
OTU6940000130k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Nectriaceae
OTU713010040k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Aspergillaceae
OTU3920000110k__Fungi
OTU666000050k__Fungi;p__Mucoromycota;c__Umbelopsidomycetes;o__GS23;f__unidentified;g__unidentified;s__unidentified
OTU5600011000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Russulales;f__Russulaceae;g__Lactarius;s__Lactarius_deceptivus
OTU37003520k__Fungi
OTU550007100k__Fungi
OTU352007000k__Fungi;p__Ascomycota;c__Dothideomycetes;o__Pleosporales
OTU493090000k__Fungi
OTU748000007k__Fungi;p__Ascomycota
OTU630050030k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Cantharellales;f__Clavulinaceae;g__Clavulina;s__Clavulina_cinerea
OTU596007000k__Fungi;p__Ascomycota
OTU3880001000k__Fungi;p__Glomeromycota
OTU670005050k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Hypocreaceae;g__Trichoderma
OTU373000260k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Boletales;f__Boletinellaceae;g__Boletinellus;s__Boletinellus_merulioides
OTU629001321k__Fungi;p__Basidiomycota;c__Cystobasidiomycetes
OTU409000040k__Fungi;p__Ascomycota;c__Dothideomycetes;o__Pleosporales
OTU489061000k__Fungi;p__Ascomycota
OTU367000700k__Fungi;p__Ascomycota
OTU662001102k__Fungi;p__Mucoromycota;c__Endogonomycetes;o__GS22;f__unidentified;g__unidentified;s__unidentified
OTU527007000k__Fungi;p__Basidiomycota;c__Tremellomycetes;o__Trichosporonales;f__Trichosporonaceae
OTU381000500k__Fungi;p__Ascomycota
OTU5620012000k__Fungi;p__Ascomycota
OTU538006000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Entolomataceae;g__Entoloma
OTU588009000k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales;f__Helotiaceae;g__Ascocoryne;s__Ascocoryne_sarcoides
OTU381140000k__Fungi
OTU428100234k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Clavicipitaceae;g__Metarhizium
OTU585003000k__Fungi;p__Ascomycota;c__Dothideomycetes
OTU87022240k__Fungi
OTU544008010k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales
OTU534004000k__Plantae;p__Anthophyta;c__Eudicotyledonae;o__Fagales;f__Fagaceae;g__Quercus;s__unidentified
OTU599005001k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Clavariaceae
OTU371000500k__Fungi;p__Ascomycota
OTU292040000k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Sordariales
OTU606006000k__Fungi
OTU609001041k__Fungi;p__Chytridiomycota;c__Rhizophydiomycetes;o__Rhizophydiales
OTU427000107k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Aspergillaceae;g__Penicillium
OTU537006000k__Fungi;p__Ascomycota
OTU600007000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Clavariaceae;g__Clavaria
OTU723000007k__Fungi;p__Ascomycota
OTU535006000k__Fungi;p__Ascomycota;c__Pezizomycetes;o__Pezizales;f__Pyronemataceae;g__Scutellinia;s__Scutellinia_cejpii
OTU389000105Unclassified
OTU722000009k__Fungi
OTU742000007k__Fungi;p__Mucoromycota;c__Umbelopsidomycetes;o__Umbelopsidales;f__Umbelopsidaceae;g__Umbelopsis;s__unidentified
OTU482310005k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Aspergillaceae;g__Penicillium
OTU416000080k__Fungi;p__Ascomycota
OTU619006010k__Fungi;p__Ascomycota
OTU519440020k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Aspergillaceae;g__Penicillium
OTU419000090k__Fungi;p__Glomeromycota;c__Archaeosporomycetes;o__Archaeosporales;f__unidentified;g__unidentified;s__unidentified
OTU603008000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Amanitaceae;g__Amanita
OTU712000070k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales
OTU553009000k__Fungi;p__Ascomycota;c__Sordariomycetes
OTU4681110001k__Fungi
OTU399000090k__Fungi;p__Basidiomycota;c__Tritirachiomycetes;o__Tritirachiales;f__Tritirachiaceae;g__Paratritirachium;s__unidentified
OTU4551020001k__Fungi
OTU232005001Unclassified
OTU281703000k__Fungi;p__Mucoromycota
OTU444803000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Sebacinales;f__Sebacinaceae;g__Sebacina;s__unidentified
OTU594003000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Clavariaceae
OTU608005000k__Fungi;p__Ascomycota
OTU356007000k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales;f__Leotiaceae;g__Leotia;s__Leotia_lubrica
OTU384001300k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella
OTU55331110k__Fungi
OTU565005010k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Clavariaceae;g__Clavulinopsis;s__Clavulinopsis_luteoalba
OTU581004000k__Fungi;p__Ascomycota;c__Lecanoromycetes;o__Lecanorales
OTU343002001k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Cortinariaceae;g__Cortinarius;s__Cortinarius_hinnuleus
OTU4471030000k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Hypocreaceae;g__Trichoderma
OTU620002000k__Fungi;p__Ascomycota
OTU702000050k__Fungi
OTU673000060k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Inocybaceae;g__Inocybe;s__Inocybe_mutata
OTU610002000k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales
OTU4432000000k__Fungi
OTU532005000k__Fungi
OTU488070000k__Fungi
OTU724000005k__Plantae;p__unidentified;c__unidentified;o__unidentified;f__unidentified;g__unidentified;s__unidentified
OTU302030000k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales;f__Helotiaceae;g__Ascocoryne
OTU50900000k__Fungi
OTU618008000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Entolomataceae;g__Entoloma;s__Entoloma_alboumbonatum
OTU156050000k__Fungi
OTU741000205k__Fungi;p__Ascomycota
OTU750000116k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella
OTU584004000k__Fungi;p__Ascomycota
OTU590004000k__Fungi;p__Glomeromycota;c__Archaeosporomycetes;o__Archaeosporales;f__Archaeosporaceae;g__Archaeospora;s__unidentified
OTU674000060k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Lycoperdaceae;g__Lycoperdon
OTU556004000k__Fungi
OTU420000060k__Fungi;p__Ascomycota
OTU700000250k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Clavariaceae;g__Camarophyllopsis;s__Camarophyllopsis_schulzeri
OTU363000500k__Fungi;p__Glomeromycota
OTU617004000k__Fungi
OTU65510000k__Fungi
OTU554002000k__Fungi;p__Chytridiomycota;c__unidentified;o__unidentified;f__unidentified;g__unidentified;s__unidentified
OTU683000030k__Fungi;p__Basidiomycota;c__Malasseziomycetes;o__Malasseziales;f__Malasseziaceae;g__Malassezia;s__Malassezia_restricta
OTU525002000k__Fungi;p__Ascomycota
OTU598004000k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales;f__Leotiaceae;g__Neobulgaria;s__Neobulgaria_pura
OTU504040000k__Fungi
OTU463305000k__Fungi;p__Ascomycota
OTU506052000k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Coniochaetales;f__Coniochaetaceae;g__Coniochaeta
OTU2781600000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Amanitaceae;g__Amanita
OTU479300010k__Fungi
OTU4391600000k__Fungi
OTU625000600k__Fungi;p__GS19;c__unidentified;o__unidentified;f__unidentified;g__unidentified;s__unidentified
OTU729000005k__Fungi;p__Ascomycota
OTU745000002k__Fungi;p__Mucoromycota;c__Mucoromycetes;o__Mucorales;f__Mucoraceae;g__Zygorhynchus;s__unidentified
OTU508050000k__Fungi;p__Ascomycota
OTU612002000k__Fungi
OTU717000040k__Fungi
OTU653000140k__Fungi;p__Ascomycota;c__Dothideomycetes;o__Pleosporales;f__Didymellaceae
OTU642000400k__Fungi;p__Ascomycota;c__Dothideomycetes;o__Pleosporales
OTU737000101k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales
OTU513032000k__Fungi;p__Ascomycota
OTU518012300k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Stachybotryaceae;g__Myrothecium;s__Myrothecium_inundatum
OTU210610000k__Fungi
OTU680000030k__Fungi;p__Ascomycota;c__Leotiomycetes
OTU499200000k__Fungi;p__Ascomycota;c__Dothideomycetes;o__Capnodiales;f__Mycosphaerellaceae
OTU86420000k__Fungi
OTU707000050k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Sordariales;f__Lasiosphaeriaceae;g__Cercophora;s__Cercophora_sparsa
OTU614004010k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella;s__Mortierella_globulifera
OTU365000300k__Fungi;p__Basidiomycota;c__Tremellomycetes;o__Trichosporonales;f__Trichosporonaceae
OTU4591001000k__Fungi;p__Ascomycota
OTU75400000k__Fungi
OTU491540000k__Plantae;p__unidentified;c__unidentified;o__unidentified;f__unidentified;g__unidentified;s__unidentified
OTU648000600k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Xylariales
OTU564004000k__Fungi
OTU4621200010k__Fungi;p__Ascomycota;c__Saccharomycetes;o__Saccharomycetales;f__Saccharomycetales_fam_Incertae_sedis;g__Candida
OTU705000010k__Fungi;p__Ascomycota
OTU696000050k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Cantharellales;f__Cantharellales_fam_Incertae_sedis;g__Sistotrema
OTU257000200k__Fungi
OTU545003000k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales;f__Helotiaceae
OTU734002002k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Hypocreales_fam_Incertae_sedis;g__Acremonium;s__Acremonium_spinosum
OTU4761200000k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales;f__Helotiales_fam_Incertae_sedis;g__Cadophora;s__Cadophora_finlandica
OTU708000050k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Clavicipitaceae;g__Pochonia;s__Pochonia_bulbillosa
OTU551004000k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Ophiocordycipitaceae;g__Tolypocladium;s__unidentified
OTU664000020k__Fungi;p__Ascomycota;c__Geoglossomycetes;o__Geoglossales;f__Geoglossaceae;g__Geoglossum;s__Geoglossum_difforme
OTU627000100k__Fungi
OTU563102010k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Coniochaetales;f__Coniochaetaceae;g__Coniochaeta
OTU651001300k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales
OTU213420000k__Fungi
OTU647000101k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Entolomataceae;g__Entoloma;s__Entoloma_terreum
OTU725100002k__Fungi;p__Ascomycota
OTU643000400k__Fungi;p__Ascomycota
OTU640003100k__Fungi;p__GS19;c__unidentified;o__unidentified;f__unidentified;g__unidentified;s__unidentified
OTU706000012k__Fungi;p__Glomeromycota;c__Archaeosporomycetes;o__Archaeosporales;f__Archaeosporaceae;g__Archaeospora;s__unidentified
OTU529004000k__Fungi;p__Ascomycota
OTU644000110k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Xylariales;f__Xylariaceae;g__Nemania;s__Nemania_serpens
OTU592002000k__Fungi;p__Ascomycota
OTU595005100k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales;f__Dermateaceae;g__Mollisia;s__unidentified
OTU661000100k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Entolomataceae;g__Entoloma;s__Entoloma_terreum
OTU744000004k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Trechisporales;f__Hydnodontaceae;g__Trechispora
OTU401000030k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales;f__Leotiaceae;g__Leotia;s__Leotia_lubrica
OTU735000001k__Fungi;p__Glomeromycota;c__Archaeosporomycetes;o__Archaeosporales;f__Archaeosporaceae;g__Archaeospora;s__unidentified
OTU718000040k__Fungi;p__Ascomycota
OTU285310111k__Fungi
OTU478400200k__Fungi
OTU709000020k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Inocybaceae;g__Inocybe;s__Inocybe_hystrix
OTU663100110k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Clavicipitaceae
OTU589004000k__Fungi;p__Ascomycota;c__Leotiomycetes;o__Helotiales;f__Hyaloscyphaceae;g__Arachnopeziza
OTU597001000k__Fungi;p__Glomeromycota;c__Archaeosporomycetes;o__Archaeosporales;f__Archaeosporaceae;g__Archaeospora;s__unidentified
OTU569300000k__Fungi
OTU583003000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Thelephorales;f__Thelephoraceae
OTU406000110k__Fungi
OTU526004000k__Fungi;p__Basidiomycota;c__Microbotryomycetes;o__Leucosporidiales;f__unidentified;g__unidentified;s__unidentified
OTU649000300k__Fungi;p__Mucoromycota;c__Umbelopsidomycetes;o__Umbelopsidales;f__Umbelopsidaceae;g__Umbelopsis;s__unidentified
OTU622000200k__Fungi;p__Ascomycota;c__Eurotiomycetes
OTU667000030k__Fungi;p__Glomeromycota;c__Glomeromycetes;o__Glomerales;f__Claroideoglomeraceae;g__Claroideoglomus
OTU287200110k__Fungi
OTU260000001k__Fungi
OTU117500000k__Fungi
OTU541002000k__Fungi;p__Glomeromycota;c__Glomeromycetes;o__Glomerales;f__Glomeraceae;g__unidentified;s__unidentified
OTU689000020Unclassified
OTU522010201k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Aspergillaceae;g__Penicillium
OTU646000010k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Boletales
OTU486710000k__Fungi;p__Basidiomycota;c__Tremellomycetes;o__Tremellales;f__Bulleraceae;g__Bullera;s__Bullera_unica
OTU568002000k__Fungi;p__Ascomycota
OTU698000010k__Fungi;p__Ascomycota
OTU449900000k__Fungi;p__Ascomycota;c__Dothideomycetes;o__Dothideomycetes_ord_Incertae_sedis;f__Gloniaceae;g__Cenococcum
OTU740002000k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Ophiocordycipitaceae;g__Haptocillium;s__Haptocillium_sinense
OTU477800000k__Fungi;p__Ascomycota
OTU465700000k__Fungi
OTU160000001k__Fungi
OTU657000101k__Fungi;p__Ascomycota
OTU631000100k__Fungi;p__Ascomycota
OTU130330000k__Fungi
OTU582002000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Cortinariaceae
OTU715000050k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Tricholomataceae;g__Tricholoma;s__Tricholoma_fulvum
OTU567021000k__Fungi;p__Ascomycota
OTU688000010k__Fungi;p__Basidiomycota;c__Tremellomycetes;o__Tremellales;f__Trimorphomycetaceae;g__Saitozyma;s__Saitozyma_podzolica
OTU502120000k__Fungi
OTU730000002k__Fungi
OTU655000100k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Hypocreales;f__Hypocreaceae;g__Trichoderma;s__Trichoderma_sinuosum
OTU471510000k__Fungi
OTU116400000k__Fungi
OTU728000002k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Polyporales;f__Coriolaceae;g__Rigidoporus;s__Rigidoporus_pouzarii
OTU691000010k__Fungi;p__Ascomycota
OTU202000001k__Fungi
OTU469400000k__Fungi;p__Mucoromycota;c__Mucoromycetes;o__Mucorales;f__Rhizopodaceae;g__Rhizopus;s__Rhizopus_arrhizus
OTU687000130k__Fungi
OTU458301000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Thelephorales;f__Thelephoraceae
OTU421000010k__Fungi;p__Ascomycota;c__Pezizomycetes;o__Pezizales;f__Pyronemataceae
OTU738000002k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Amanitaceae;g__Amanita
OTU369000200k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Atheliales;f__Atheliaceae;g__Athelia
OTU676000020k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Clavariaceae;g__Clavaria
OTU364000200k__Fungi;p__Chytridiomycota;c__Rhizophydiomycetes;o__Rhizophydiales;f__Rhizophydiaceae;g__Rhizophydium;s__unidentified
OTU637000201k__Fungi
OTU711000020k__Fungi;p__Ascomycota
OTU484200001k__Fungi;p__Ascomycota;c__Saccharomycetes;o__Saccharomycetales;f__Pichiaceae;g__Pichia;s__Pichia_kluyveri
OTU660000300k__Fungi;p__Ascomycota
OTU601002000k__Fungi;p__Basidiomycota;c__Agaricomycetes
OTU695000010k__Fungi;p__Basidiomycota
OTU326003000k__Fungi;p__Ascomycota;c__Sordariomycetes
OTU615001000k__Fungi
OTU511200000k__Fungi
OTU686001010k__Fungi;p__Ascomycota
OTU497020000k__Fungi
OTU633010000k__Fungi
OTU624000101k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Chaetosphaeriales;f__Chaetosphaeriaceae
OTU645000100k__Fungi
OTU483320000k__Fungi
OTU386000200k__Fungi
OTU457510000k__Fungi
OTU214300000k__Fungi
OTU472300000k__Fungi
OTU277310000k__Fungi
OTU693000010k__Plantae;p__unidentified;c__unidentified;o__unidentified;f__unidentified;g__unidentified;s__unidentified
OTU425000010k__Fungi;p__Ascomycota;c__Eurotiomycetes;o__Eurotiales;f__Aspergillaceae
OTU297020000k__Fungi
OTU611001000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Clavariaceae
OTU212200001k__Fungi
OTU448500000k__Fungi;p__Basidiomycota;c__Tremellomycetes;o__Filobasidiales;f__Piskurozymaceae;g__Solicoccozyma;s__Solicoccozyma_terrea
OTU512110010k__Fungi
OTU586003000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Trechisporales;f__Hydnodontaceae;g__Brevicellicium;s__Brevicellicium_exile
OTU496020000k__Fungi;p__Ascomycota;c__Dothideomycetes;o__Dothideomycetidae_ord_Incertae_sedis;f__Eremomycetaceae;g__Arthrographis;s__unidentified
OTU703000030k__Fungi;p__Chytridiomycota;c__Chytridiomycetes;o__Chytridiales;f__Chytriomycetaceae
OTU451220000k__Fungi
OTU654000100k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Glomerellales;f__Plectosphaerellaceae;g__Plectosphaerella
OTU208210010k__Fungi
OTU498010000k__Fungi
OTU288300000k__Fungi
OTU507030000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Polyporales;f__Coriolaceae
OTU523120000k__Fungi
OTU679000030k__Fungi;p__Ascomycota
OTU296010000k__Fungi;p__Basidiomycota;c__Agaricomycetes
OTU291020000k__Fungi
OTU286200000k__Fungi
OTU656000100k__Fungi;p__Ascomycota;c__Pezizomycetes;o__Pezizales;f__Pyronemataceae;g__Sphaerosporella;s__unidentified
OTU714000020k__Fungi;p__Ascomycota;c__Sordariomycetes
OTU467300000k__Fungi
OTU659000300k__Fungi
OTU283300000k__Fungi
OTU571001000k__Fungi;p__Basidiomycota
OTU480400000k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Atheliales;f__Atheliaceae;g__Piloderma;s__unidentified
OTU747000001k__Fungi;p__Basidiomycota;c__Agaricomycetes;o__Agaricales;f__Entolomataceae;g__Entoloma;s__unidentified
OTU279400000k__Fungi
OTU470300000k__Fungi
OTU746000002k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Chaetosphaeriales;f__Chaetosphaeriaceae;g__Chaetosphaeria
OTU501010000k__Fungi
OTU500010000k__Fungi;p__Ascomycota;c__Sordariomycetes
OTU473200000k__Fungi
OTU481300000k__Fungi
OTU460300000k__Fungi
OTU453300000k__Fungi
OTU475200000k__Fungi
OTU716000020k__Fungi;p__Ascomycota
OTU616002000k__Fungi;p__Ascomycota;c__Leotiomycetes
OTU490010000k__Fungi
OTU503010000k__Fungi
OTU579001001k__Fungi
OTU461300000k__Fungi
OTU675000020k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Diaporthales;f__Valsaceae
OTU452300000k__Fungi;p__Ascomycota;c__Sordariomycetes;o__Sordariales;f__Cephalothecaceae;g__Phialemonium
OTU510020000k__Fungi
OTU668000010k__Fungi
OTU638000200k__Fungi
OTU474200000k__Fungi
OTU515010000k__Fungi
OTU454300000k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella;s__Mortierella_pseudozygospora
OTU720000001k__Fungi;p__Mortierellomycota;c__Mortierellomycetes;o__Mortierellales;f__Mortierellaceae;g__Mortierella;s__Mortierella_echinula

3.3.2. OTU heatmap

The heatmap analysis shows the abundance information of selected OTU as well as the similarity and difference across OTUs and samples by similarity clustering. The figure below shows the top 30 OTUs with the highest abundance:

Figure 3.3.2 OTU abundance clustering heatmap

Note: The row name is the OTU ID, the column name is the sample information, the left side of the figure is the OTU cluster tree, and the top is the sample cluster tree. The value of each colored box is the relative abundance of each OTU after normalization.

3.3.3. OTU Venn diagram and petal diagram

According to the results of OTU cluster analysis, the common and unique OTUs of different samples/groups are analyzed. When the number of samples/groups is less than 5, the Venn diagram is drawn. When the sample/group is greater than 5, the petal diagram is drawn.

result/01_OTU_Taxa/OTU/Venn/venn.png
  • result/01_OTU_Taxa/OTU/Venn/venn.png

Figure 3.3.3 OTU Venn diagram or petal diagram

Note: the circles of different colors in the Venn diagram represent different samples or groups, and the numbers in the figure represent the numbers of OTUs unique or common to each sample or group. In the petal diagram, each petal represents a sample or group. The numbers on the petals represent the number of OTUs unique to the sample, and the white circle in the middle represents the number of OTUs shared by all samples and groups.

3.3.4. Species annotation statistics

In order to obtain the classification information of OTU, a representative sequence was selected for each OTU and annotated using the RDP classifier, thereby to obtain the community composition of each sample.

Analysis software: Qiime(1.9.1)

Analysis method: RDP classifier Bayesian algorithm was used to classify the OTU representative sequences of 97% similarity level, and the community composition of each sample was analyzed and summarized at all levels. The comparison database was Greengenes database (http://qiime.org/home_static/dataFiles.html) / 18S rRNA database / ITS database.

For each sample, the percentage of each species at different taxonomic levels (Phylum, Class, Order, Families, Genus, Species) is shown in the table below:

TaxonC1C2-2C3P1P2P3
Unclassified4,7226,55017,2837,9598,9257,270
Mucor18,074.0010,057.00196.00329.00231.001.00
Russula368.00552.002,438.001,046.002,873.0010,531.00
Sebacina17.000.00147.003,934.002,582.003,704.00
Pseudotricholoma45.009,258.00161.001.002.000.00
Saitozyma214.0082.00532.003,803.002,936.001,550.00
Leohumicola781.00456.00328.00450.001,876.001,865.00
Elaphomyces3,600.002.004.002,095.003.002.00
Geminibasidium63.00108.001,592.003,086.00478.0025.00
Sagenomella26.000.00100.00173.003,509.00674.00
Lactarius0.000.00443.002,710.0099.00297.00
Scleroderma0.000.0017.007.002,465.003.00
Mortierella18.0080.001,004.00255.00299.00308.00
Astraeus0.000.004.001,097.00346.00130.00
Penicillium29.0056.00365.00149.00300.00268.00
Trichoglossum4.00940.000.001.001.0011.00
Cenococcum192.0044.00452.007.00198.007.00
Clavulinopsis3.000.00689.002.006.002.00
Trichoderma11.0017.00306.00166.0089.0030.00
Inocybe0.000.0015.00310.00252.002.00
Pachyphloeus0.000.001.002.00293.00236.00
Clavaria1.00199.00272.0016.004.0032.00
Hydnobolites0.000.00405.000.0014.000.00
Tuber0.007.00344.001.001.000.00
Tylopilus0.000.000.00172.000.00171.00
Amanita18.000.00144.001.002.00171.00
Umbelopsis8.0018.0057.0017.0095.00134.00
Ilyonectria3.003.00112.00115.0088.008.00
Chaetosphaeria0.000.0034.002.00118.00168.00
Bondarzewia9.003.0013.00155.00135.005.00
Trichocladium25.000.0032.0056.0030.00105.00
Cuphophyllus0.00120.00125.000.001.000.00
Metarhizium1.0025.0035.0017.0073.0089.00
Tomentella0.0032.009.0099.000.0099.00
Tricholoma213.000.000.000.005.000.00
Pseudogymnoascus2.007.0021.0036.0038.00104.00
Byssocorticium0.000.00202.000.003.000.00
Rhizophagus1.000.0013.0011.000.00174.00
Gliocladiopsis0.000.000.001.000.00148.00
Cudoniella0.000.000.00138.000.000.00
Solicoccozyma6.001.0052.0027.008.0029.00
Dendrosporium9.009.0019.0038.003.0039.00
Calostoma0.000.00115.001.000.000.00
Lactifluus0.000.000.001.001.00101.00
Pseudoclathrosphaerina0.000.000.0040.0053.005.00
Leptodontidium2.005.0032.0012.005.0042.00
Pseudotomentella0.000.0064.000.002.000.00
Chaetomium2.001.000.0029.0017.0016.00
Cortinarius1.000.0061.000.001.001.00
Camarophyllopsis0.000.0047.002.005.000.00
Neoascochyta46.002.000.003.001.002.00
Dioszegia47.002.001.003.000.000.00
Entoloma0.000.0044.002.000.002.00
Genea0.000.0045.000.000.000.00
Archaeospora0.000.005.000.009.0029.00
Mycena35.003.000.000.000.000.00
Clonostachys0.000.000.002.0026.0010.00
Lycoperdon0.000.0026.000.006.005.00
Ceratobasidium0.000.000.000.0031.000.00
Delicatula3.000.003.000.0023.000.00
Rhizodermea15.000.0011.000.001.000.00
Dissophora0.000.0025.000.000.000.00
Asterophora0.000.0024.000.000.000.00
Nectria0.000.000.0023.000.000.00
Rectipilus8.000.0014.000.000.000.00
Coniochaeta1.005.004.0010.001.000.00
Ramariopsis0.000.0019.000.000.000.00
Trechispora0.000.0014.000.000.005.00
Ambispora0.000.0019.000.000.000.00
Nothomitra0.000.0018.000.000.000.00
Tremella0.000.000.000.000.0018.00
Volutella0.000.002.0016.000.000.00
Malassezia0.007.000.000.0010.000.00
Ramularia0.000.000.003.009.004.00
Cyberlindnera0.000.000.000.000.0015.00
Gliophorus0.000.0015.000.000.000.00
Laccaria0.000.000.000.0015.000.00
Acremonium0.000.002.000.000.0013.00
Leucothecium0.002.001.002.007.002.00
Fenestella0.000.000.0014.000.000.00
Ganoderma0.001.000.000.0013.000.00
Pestalotiopsis0.000.0014.000.000.000.00
Strobilomyces0.000.0013.000.000.000.00
Candida12.000.000.000.001.000.00
Cadophora12.000.000.000.000.000.00
Ascocoryne0.003.009.000.000.000.00
Hydnotrya0.000.0011.000.000.000.00
Leotia0.000.007.000.003.000.00
Sistotrema0.000.000.005.005.000.00
Paratritirachium0.000.000.000.009.000.00
Boletinellus0.000.000.002.006.000.00
Bullera7.001.000.000.000.000.00
Clavulina0.005.000.000.003.000.00
Scutellinia0.000.006.000.000.000.00
Talaromyces0.001.002.002.001.000.00
Myrothecium0.001.002.003.000.000.00
Mollisia0.000.005.001.000.000.00
Leucogaster0.000.000.000.006.000.00
Cercophora0.000.000.000.005.000.00
Pochonia0.000.000.000.005.000.00
Quercus0.000.004.000.000.000.00
Piloderma4.000.000.000.000.000.00
Arachnopeziza0.000.004.000.000.000.00
Tolypocladium0.000.004.000.000.000.00
Neobulgaria0.000.004.000.000.000.00
Rhizopus4.000.000.000.000.000.00
Brevicellicium0.000.003.000.000.000.00
Phialemonium3.000.000.000.000.000.00
Pichia2.000.000.000.000.001.00
Claroideoglomus0.000.000.000.003.000.00
Nemania0.000.000.001.001.000.00
Geoglossum0.000.000.000.002.000.00
Rhizophydium0.000.000.002.000.000.00
Arthrographis0.002.000.000.000.000.00
Rigidoporus0.000.000.000.000.002.00
Athelia0.000.000.002.000.000.00
Zygorhynchus0.000.000.000.000.002.00
Haptocillium0.000.002.000.000.000.00
Plectosphaerella0.000.000.001.000.000.00
Sphaerosporella0.000.000.001.000.000.00

Table 3.3.4.1 Taxa Statistics at Genus level

SamplesC1C2-2C3P1P2P3
Kingdom223333
Phylum899989
Class182026222624
Order303144424238
Family404060566350
Genus453976647054
Species373669576552

Table 3.3.4.2 Statistics of Taxonomic Composition

3.3.5. Graphics display of species relative abundance

The distribution of the top 30 most abundant classifications in each sample or group at different taxonomic levels (Phylum, Class, Order, Families, Genus, Species) are shown as follows:

result/01_OTU_Taxa/Taxonomy/Community/Family_abundance_community.png
  • result/01_OTU_Taxa/Taxonomy/Community/Family_abundance_community.png
  • result/01_OTU_Taxa/Taxonomy/Community/Genus_abundance_community_Group.png
  • result/01_OTU_Taxa/Taxonomy/Community/Class_abundance_community_Group.png
  • result/01_OTU_Taxa/Taxonomy/Community/Phylum_abundance_community.png
  • result/01_OTU_Taxa/Taxonomy/Community/Species_abundance_community_Group.png
  • result/01_OTU_Taxa/Taxonomy/Community/Phylum_abundance_community_Group.png
  • result/01_OTU_Taxa/Taxonomy/Community/Species_abundance_community.png
  • result/01_OTU_Taxa/Taxonomy/Community/Order_abundance_community_Group.png
  • result/01_OTU_Taxa/Taxonomy/Community/Family_abundance_community_Group.png
  • result/01_OTU_Taxa/Taxonomy/Community/Genus_abundance_community.png
  • result/01_OTU_Taxa/Taxonomy/Community/Class_abundance_community.png
  • result/01_OTU_Taxa/Taxonomy/Community/Order_abundance_community.png

Figure 3.3.5.1 Stacked bar plot of species distribution

Note: X axis is the sample name or group name, and the Y axis is the relative abundance of different species. The legend is the name of the taxonomic classification of the species. 'Other' represents the relative abundance of all phylum level classifications other than the top 30.

The top 30 species distribution of each sample (or group) on different levels (Phylum, Class, Order, Families, Genus, Species) was clustered and plotted in a heatmap. The similarity and difference of each species is visualized by color scheme in the heat map. The heat maps plotted for the distribution of the species in each sample on different levels of classification are shown below:

result/01_OTU_Taxa/Taxonomy/Heatmap/Class_abundance_heatmap.png
  • result/01_OTU_Taxa/Taxonomy/Heatmap/Class_abundance_heatmap.png
  • result/01_OTU_Taxa/Taxonomy/Heatmap/Order_abundance_heatmap.png
  • result/01_OTU_Taxa/Taxonomy/Heatmap/Phylum_abundance_heatmap.png
  • result/01_OTU_Taxa/Taxonomy/Heatmap/Species_abundance_heatmap.png
  • result/01_OTU_Taxa/Taxonomy/Heatmap/Family_abundance_heatmap.png
  • result/01_OTU_Taxa/Taxonomy/Heatmap/Family_abundance_Group.png
  • result/01_OTU_Taxa/Taxonomy/Heatmap/Genus_abundance_heatmap.png
  • result/01_OTU_Taxa/Taxonomy/Heatmap/Species_abundance_Group.png
  • result/01_OTU_Taxa/Taxonomy/Heatmap/Class_abundance_Group.png
  • result/01_OTU_Taxa/Taxonomy/Heatmap/Genus_abundance_Group.png
  • result/01_OTU_Taxa/Taxonomy/Heatmap/Phylum_abundance_Group.png
  • result/01_OTU_Taxa/Taxonomy/Heatmap/Order_abundance_Group.png

Figure 3.3.5.2 Species distribution heatmap

Note: The columns represent samples and/or groups and the rows represent species. The dendrogram above the heatmap is the cluster result of the samples and/or groups and the dendrogram to the left is the species cluster. The colors in the heat map represent the relative abundance of the corresponding species in the corresponding sample or group.

3.4. Sample complexity analysis

3.4.1. Alpha diversity analysis

In community ecology, alpha diversity is mainly used to reflect the diversity of each sample, which estimates the number of species in the microbial community as well as the abundance and diversity of species in environmental communities through a series of statistical indices.

The indices for community richness calculation include:

ACE: An index to estimate the number OTU in communities. It was proposed by the Chao and is commonly used to estimate the total number of species in ecology. ( http://www.mothur.org/wiki/Ace )

Chao: It is an index that uses the Chao 1 algorithm to estimate the OTU number of samples. Chao is commonly used in ecology to assess the total number of microorganisms.( http://www.mothur.org/wiki/Chao )

The indices for community diversity calculation include:

Shannon: Commonly used to reflect the diversity index a for the estimation of microbial diversity. ( http://www.mothur.org/wiki/Shannon )

Simpson: Simpson diversity index, proposed by Edward Hugh Simpson in 1949 and is commonly used in ecology to quantify biological diversity in a region. ( http://www.mothur.org/wiki/Simpson )

The indices to measure sequencing depth (Coverage) include:

Goods Coverage: Refers to library coverage of each sample. The higher the value, the lower the probability that the sample did not cover the sequence.(http://www.mothur.org/wiki/Coverage )

Analysis software:Qiime (1.9.1)

Analysis method: sequences were randomly extracted and the valid sequences were subject to OTU analysis and a diversity index was calculated for each sample.

Alpha diversity results are summarized in the table below:

sampleacechao1shannonsimpsongoods_coverage
C1215.442142.460.591.00
C2-2218.14217.733.080.771.00
C3407.14406.836.340.971.00
P1343.04332.174.830.931.00
P2416.81412.775.020.951.00
P3323.86344.754.800.901.00

Table 3.4.1 Collation of alpha diversity results

3.4.2. Between-group differential analysis using alpha-diversity indices

To perform between-group a diversity analysis, box plots were generated based on a diversity indices using R, which intuitively displays the maximum, minimum, median and outliers of the a diversity indices of samples in each group as well as the differences between groups.

Analysis method: Box plot was generated using R based on a-diversity index.

The box plot of the inter-group differential analysis based on chao1 and shannon indices is as below:

result/02_Alpha_Diversity/Alpha_Index/chao1.png
  • result/02_Alpha_Diversity/Alpha_Index/chao1.png
  • result/02_Alpha_Diversity/Alpha_Index/shannon.png

Note: The left panel is the Chao1 index boxplot of each group. X axis indicates the names of the groups and Y axis indicates the Chao 1 index. Each box diagram shows the minimum, first quartile, medium, third quartile and maximum values of the chao1 index of the corresponding sample. The right graph is the Shannon index boxplot of each group.

3.4.3. Rank-abundance curve

Rank-abundance curve is used to analyze diversity. To generate a rank-abundance curve, the number of valid sequences in each OTU of a given sample was first calculated, and then all the OTUs were ranked in descending order based on their relative abundance (number of valid sequences), and finally the result was plotted with OTU ranking on the X axis and the number of sequences in the OTU on the Y axis. Y axis could also be OTU relative abundance in percentage.

Rank-abundance curve reflects both species abundance and species uniformity. The abundance of species is reflected by the length of the curve on the X axis. The more extended on the X axis, the more abundant the species is. Species uniformity is reflected by the shape of the curve. The smoother the curve, the higher the species uniformity.

Analysis method: R packages were used for graph generation based on the results of OTU analysis

Figure 3.4.3 Rank-Abundance curve

Note: Each curve in the figure above corresponds to an individual sample. The X axis is the relative abundance of the OTU in descending order. The Y axis is the relative abundance of the OTU. '100' on the X axis indicates the OTU in the sample is ranked as the 100th abundant in descending order, and the corresponding value on the Y axis is the percentage of the sequence count in the OTU (the number of sequences of the OTU divided by the total number of sequences).

3.4.4. Rarefaction curve

The rarefaction curve is a useful tool to characterize the species composition of a sample and predicting the abundance of species in a sample. It efficiently deals with the increase of detected species due to the increase in sample size. It is widely used in biodiversity and community surveys to determine whether the sample size is sufficient and to estimate the species abundance. Therefore, the rarefaction curve can not only determine whether the sample size is sufficient, but also predict the species abundance when the sample size is sufficient.

Analysis software: Qiime (1.9.1)

Analysis method: The rarefaction curve was constructed by random sampling. The observed numbers of OTUs were plotted against the number of extracted sequences.

Figure 3.4.4 Observed OTUs rarefaction curves

Note: The X axis is the number of valid sequences extracted, and the Y axis is the number of OTUs (Observed OTUs). Each sample is represented by one curve with a unique color. The number of OTUs increases with the increase of extracted sequence count until reaching a plateau, which indicates the number of detected OTUs will not increase with the amount of extracted sequences and reflects the reasonable sequence depth.

3.5. Multi-sample comparative analysis

3.5.1. (Un)weighted unifrac distance matrix

Beta diversity reflects the diversity and the degrees of differences among samples. The distance between the samples can be calculated using the evolution and abundance information between the sample sequences to reflect whether there is significant difference in microbial community among the samples. This can be achieved by UniFrac analysis.

Analysis software: Qiime (1.9.1)

Analysis method: a phylogenetic tree was constructed using the OTU representative sequences from different environmental samples. The Unifrac metric was then used to measure the difference between two different environmental samples according to the length of the constructed evolutionary tree.

UniFrac analysis includes weighted unifrac and unweighted unifrac methods. The difference between the two is whether to include the relative abundance of sequences from different environmental samples. The weighted unifrac algorithm weights the sequence abundance information when calculating the length of the branch, so unweighted unifrac detects the change among the samples, and the weighted unifrac further quantifies the variation on different pedigrees.

result/03_Beta_Diversity/Unifrac_distance//Heatmap_weighted_unifrac.png
  • result/03_Beta_Diversity/Unifrac_distance//Heatmap_weighted_unifrac.png
  • result/03_Beta_Diversity/Unifrac_distance//Heatmap_unweighted_unifrac.png

Figure 3.5.1 (Un)weighted unifrac distance matrix heatmap

Note: (Un)weighted unifrac distance matrix heat map. The color scheme in the heatmap represents the degree of difference between the two samples. The lighter the color, the smaller the coefficient between the two samples, and the smaller the difference of species diversity.

3.5.2. PCoA analysis

PCoA (Principal Co-ordinates Analysis) analysis characterizes and visualizes the similarities and differences of data. Similar to PCA, it sorts data based on a series of eigenvalues and eigenvectors and uses the top ranked eigenvalues to determine the most important coordinates in the distance matrix. It also uses eigenanalysis to perform a rigid rotation of the original axes that only changes the coordinates but not the positional relationship of different sample points. The difference between the two is PCA determines the principal components based on sample similarity coefficient matrix whereas PCoA uses distance matrix to find the primary coordinates.

Analysis software: R

Analysis method: Analysis method: PCoA analysis was performed and plotted based on Brary-Curtis distance matrix

result/03_Beta_Diversity/PCoA/PC1_vs_PC3_group_plots.Group.png
  • result/03_Beta_Diversity/PCoA/PC1_vs_PC3_group_plots.Group.png
  • result/03_Beta_Diversity/PCoA/PC2_vs_PC3_plots.png
  • result/03_Beta_Diversity/PCoA/PC2_vs_PC3_group_plots.Group.png
  • result/03_Beta_Diversity/PCoA/PC1_vs_PC2_group_plots.Group.png
  • result/03_Beta_Diversity/PCoA/PC1_vs_PC3_plots.png
  • result/03_Beta_Diversity/PCoA/PC1_vs_PC2_plots.png

Figure 3.5.2 PCoA Plot

Note: Samples of the same group are represented in the same color and shape. PC1_vs_PC2 is the PCoA plot obtained for the first and second principal coordinates; the X and Y axes represent the first and second principal coordinates, respectively. The value in percentage in the axis label represents the contribution of the corresponding coordinate to the sample variance and measures how much this principal is extracted from the original information. The distance between the sample points indicates the similarity of the microbial community in the sample. The closer the points, the higher the similarity. Samples clustered together are composed of similar microbial compositions.

3.5.3. PCA analysis

PCA analysis (Principal Component Analysis) is a statistical technique for the determination of the key variables in a multidimensional data set that are most responsible for the differences in the observations, and thus is commonly used to simplify complex data analysis.

The difference and distance between samples can be reflected by the analysis of the gene functional distribution of different samples. The differences between multiple sets of data can be plotted on a two-dimensional chart using variance decomposition, with the axes representing two eigenvalues that reflects the largest variance values.

result/03_Beta_Diversity/PCA/PC1_vs_PC3_group_plots.Group.png
  • result/03_Beta_Diversity/PCA/PC1_vs_PC3_group_plots.Group.png
  • result/03_Beta_Diversity/PCA/PC2_vs_PC3_plots.png
  • result/03_Beta_Diversity/PCA/PC2_vs_PC3_group_plots.Group.png
  • result/03_Beta_Diversity/PCA/PC1_vs_PC2_group_plots.Group.png
  • result/03_Beta_Diversity/PCA/PC1_vs_PC3_plots.png
  • result/03_Beta_Diversity/PCA/PC1_vs_PC2_plots.png

Figure 3.5.3 PCA graph

Note: PC1, PC2, PC3 represent the first, second and third principal components, respectively. The percentage after the principal component represents the contribution rate of this component to sample difference and measures how much information the principal component can extract from the original data. The distance between samples indicates the similarity of the distribution of functional classifications in the sample. The closer the distance, the higher the similarity.

3.5.4. NMDS analysis

The non-metric multidimensional scaling is a data analysis method that reduces multi-dimensional space to low-dimensional space to simplify the localization, analysis and classification of research objects. This method preserves the primitive relation among the objects. Its main feature is to position each object in multi-dimensional space based on its functional classification information and calculate the distances between different objects (points) as a measurement of their difference, which are used to obtain the spatial position map.

Analysis method: Graph was generated using vegan package in R based on the beta diversity distance matrix.

result/03_Beta_Diversity/NMDS/NMDS.png
  • result/03_Beta_Diversity/NMDS/NMDS.png
  • result/03_Beta_Diversity/NMDS/Group_NMDS.png

Figure 3.5.4 NMDS plot

Note: Each point represents a sample, and the distance between the points indicates the degree of difference. Samples of the same group are represented by the same color. Stress < 0.2 indicates NMDS can accurately reflect the difference between the samples.

3.5.5. UPGMA Tree

Cluster analysis uses evolutionary information derived from sample sequences to calculate whether samples in a specific environment is significantly different from a evolutionary lineage in microbial communities.

Analysis software: Qiime (1.9.1)

Analysis method:The UPGMA (Unweighted pair group method with arithmetic mean) clustering method was used to cluster the samples based on the Brary-Curtis distance matrices.

result/03_Beta_Diversity/UPGMA_Tree/UPGMA_Tree_Group.png
  • result/03_Beta_Diversity/UPGMA_Tree/UPGMA_Tree_Group.png
  • result/03_Beta_Diversity/UPGMA_Tree/UPGMA_Tree.png

Figure 3.5.5 UPGMA tree

Note: Each branch in the figure represents a sample. Different colors representing different groups.

3.6. Differential analysis of community composition

3.6.1. Anosim analysis

ANOSIM is a nonparametric test used to examine whether the difference between two or more groups is statistically greater than the within-group difference. It was used to determine whether the grouping is meaningful. R value was obtained by analyzing the sample distance matrix.

Analysis software: vegan package in R

FactorR-ValueP-Value
Group0.780.10

Table 3.6.1 Group difference evaluation by Anosim

Column description:

(1) Factor:The factors used for grouping

(2) R value: The value of R ranges from 0 to 1. The closer it is to 0, the less significant the between-group difference is compared to within-group difference; the closer it is to 1, the more significance the between-group difference compared to within-group difference.

(3) P value: P-value indicates the statistical credibility, P < 0.05 indicates the result is statistically significant.

The results of the Anosim analysis were ranked by the value of the distances between the two samples. For each group pair, three set of data can be obtained: between-group distances and within-group distances of each of the two groups. The result is shown below in box plot.

result/04_Diff_Comparison/Anosim/AnosimGroup.png
  • result/04_Diff_Comparison/Anosim/AnosimGroup.png

Figure 3.6.1 Group comparison by Anosim analysis

Note: X axis is the ranking of the distances of samples. 'Between' represents the result between the two groups, the other two are the results of within-group distance calculation. R values close to 1 indicates the difference between the groups is significantly greater than the within-group difference. P < 0.05 indicates the result is statistically significant.

4. Deliverables

00_Data: Raw data statistics

01_OTU_Taxa: OTU and Taxonomy results

02_Alpha_Diversity: Sample conplexity analysis results

03_Beta_Diversity: Multi-sample comparative analysis results

04_Diff_Comparison: Differential analysis results of community composition

5. Reference

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[2] Crawford, P. A., Crowley, J. R., Sambandam, N., Muegge, B. D., Costello, E. K., Hamady, M., et al. (2009). Regulation of myocardial ketone body metabolism by the gut microbiota during nutrient deprivation. Proc Natl Acad Sci U S A, 106(27), 11276-11281.

[3] DeSantis TZ, Hugenholtz P, Larsen N, Rojas M, Brodie EL, et al. (2006) Greengenes, a chimera-checked 16S rRNA gene database and workbench compatible with ARB. Appl Environ Microbiol 72: 5069-5072 doi:10.1128/AEM.03006-05.

[4] Klindworth A, Pruesse E, Schweer T, Peplies J, Quast C, Horn M, Glockner FO (2013) Evaluation of general 16S ribosomal RNA gene PCR primers for classical and next-generation sequencing-based diversity studies. Opens external link in new windowNucl. Acids Res. 41:e1

[5] Westram R, Bader K, Pruesse E, Kumar Y, Meier H, Glockner FO, Ludwig W (2011) ARB: a software environment for sequence data. In: de Bruijn FJ (ed) Handbook of Molecular Microbial Ecology I: Metagenomics and Complementary Approaches.Opens external link in new window John Wiley & Sons, Inc., pp 399-406

[6] Yu Wang, Hua-Fang Sheng, et al. Comparison of the Levels of Bacterial Diversity in Freshwater, Intertidal Wetland, and Marine Sediments by Using Millions of Illumina Tags. Appl. Environ. Microbiol. 2012, 78(23):8264. DOI: 10.1128/AEM.01821-12.8

[7] Price MN, Dehal PS, Arkin AP (2010) FastTree 2 - Approximately Maximum-Likelihood Trees for Large Alignments. PLoS ONE 5(3): e9490. doi:10.1371.journal.pone.0009490.

[8] Micah Hamady, Catherine Lozupone and Rob Knight. Fast UniFrac: facilitating high-throughput phylogenetic analyses of microbial communities including analysis of pyrosequencing and PhyloChip data. The ISME Journal (2010) 4, 17-27; doi:10.1038/ismej.2009.97